Proposal Summary Stemness and Differentiation Potential Are Defined by Metabolism in Cardiac Progenitor Cells The limited capacity for regeneration in the adult mammalian heart can be augmented by c-Kit+ cardiac progenitor cells (CPCs). However, reparative potential of CPCs is compromised by genetic and environmental factors including aging and pathological injury. Similarly, genetic and environmental factors influence the metabolic profile and mitochondrial function of CPCs isolated from adult human heart tissue. Moreover, isolation and expansion of CPCs activates metabolic pathways that are dormant in quiescent cells. Several studies have established a link between metabolism and stem cell phenotypic characteristics of pluripotency, commitment, and viability. For example, glycolysis supports rapid cell proliferation while minimizing the production of damaging reactive oxygen species. In contrast, mitochondrial respiration is essential for differentiation, but oxidative metabolism depletes the stem cell pool and accelerates cellular senescence. The hypothesis of this project is that metabolism and stemness of human CPCs are interrelated such that CPC stemness is dependent on a glycolytic phenotype and differentiation requires biogenesis of mitochondria for oxidative metabolism. To address this hypothesis, two specific aims are proposed: (1) Metabolism regulates CPC biology: glycolysis augments proliferation and delays senescence, while oxidative phosphorylation enhances lineage commitment; and (2) Predisposition to lineage commitment can be augmented by increasing mitochondrial function. The significance of this research lies in applying findings to predict CPC function derived from patient samples to optimize regenerative treatment for individuals based on stem cell metabolic phenotype. The short-term goal of the project is to optimize in vitro expansion and lineage commitment of human CPCs by modulating metabolism. The long-term goal is the identification of drug-targetable regulators of metabolism to improve the regenerative capacity of CPCs in patients. The novelty of this research rests in elucidating relationships between metabolism, self-renewal and lineage commitment in human CPCs. The impact of this research will be to reveal how stress conditions and changes in microenvironment affect the metabolic phenotype of cardiac stem cells, thereby establishing new interventional approaches and potentiate the likelihood of success for CPC-based interventional therapies.